Neuronal death is a prominent event during normal development of animals and in may human neurological disorders. Immunochemical markers for dying neurons would be valuable reagents for they would allow a simpler, more direct way to identify dying neurons than is currently available. In the future they might also be developed as a diagnostic tool to detect neuronal death from samples of bodily fluids. The specific aims of this project are to obtain detailed data about the cytological features and time course of neuronal death after neuronal axotomy in the rat. Using that information monoclonal antibodies will be developed that specifically identify dying central and peripheral neurons. Spinal cord and ganglion will be examined histologically between 1 and 50 days after unilateral neonatal axotomy in rats. That procedure results in a massive depletion of motor neurons and ganglionic neurons. Comparisons will be made between the experimental and sham-operated sides of each rat to identify the cytological features of those neurons stained with dyes that have also ceased to incorporate tritiated leucine into protein as determined autoradiographically. Markedly diminished protein synthesis in neuronal somata is an early characteristic of neuronal death. The time course of neuronal death will be assessed directly from those tissue sections by counting dying neurons and, independently in a separate series of rats, by counting surviving, retrogradely labeled neurons. Mice will be immunized with either spinal cord or ganglionic tissue obtained from rats at a time when neuronal death is maximal after neonatal axotomy. Lymphocytes from the immunized mice will be fused with myeloma cells to make hybridomas which will be grown in tissue culture medium selective for the hybrid cells. Supernatants which contain primary antibody will be assayed on tissue sections in conjunction with a second antibody that will contain a marker. Hybridomas that produce antibodies which give a positive reaction for dying neurons relative to controls will be isolated by limiting-dilution to obtain monoclonal antibodies.